PI3K/Akt-dependent transcriptional regulation and activation of BMP-2-Smad signaling by NF-kappaB in metastatic prostate cancer cells.
BACKGROUND
Bone morphogenetic proteins (BMPs) exert osteoinductive effects in prostate cancer (PC) via uncharacterized mechanisms. In this study, we investigated whether the nuclear transcription factor NF-kappaB, implicated in PC metastasis, is involved in transcriptional regulation and activation of BMP-2 or BMP-4/Smad signaling in PC cells.


METHODS
NF-kappaB inhibition was achieved by IkappaBalpha super-repressor adenoviral vector and activation was monitored by EMSA and reporter assays. BMP expression and activation was measured by PCR and reporter assays. Promoter binding assay was performed by chromatin immunoprecipitation (ChIP) assay. Smad1/5/8 phosphorylation was measured by Western blot analysis.


RESULTS
PCR and chimeric BMP-2 and BMP-4 luciferase assays demonstrate that NF-kappaB confers robust and selective activation of BMP-2 in p65 overexpressing or rhTNF-alpha-stimulated PC cells. Inhibition of NF-kappaB significantly reduced transcript levels and autocrine production of BMP-2 by rhTNF-alpha stimulated C4-2B cells and to a lesser extent by the parental LNCaP cells. Selective inhibition of PI3K/Akt suppressed the NF-kappaB-induced BMP-2 promoter activity. Furthermore, suppression of NF-kappaB activation decreased the transcript levels and BMP-2-induced phosphorylation of Smad1/5/8, critical downstream targets of BMP-2 signaling in PC cells. Notably, the activation of BMPRII by BMP-2 is required for modulation of Smad activation by NF-kappaB in PC cells. Based on ChIP analysis, the transcriptional regulation of BMP-2 gene by NF-kappaB may be partially attributed to binding to kappab site on the BMP-2 promoter.


CONCLUSIONS
The data suggest that PI3K/Akt-NF-kappaB axis may promote PC bone metastasis in part by regulating transcription and activation of the BMP-2-Smad signaling cascade in osteotropic PC cells.
Graham
Tisheeka R
TR
Department of Urology, Tulane University Health Sciences Center, New Orleans, Louisiana 70112, USA.
Odero-Marah
Valerie A
VA
Chung
Leland W
LW
Agrawal
Krishna C
KC
Davis
Rodney
R
Abdel-Mageed
Asim B
AB
D055396:Bone Morphogenetic Protein 2 / Q000235:genetics / Q000502:physiology*; D055415:Bone Morphogenetic Protein 4 / Q000235:genetics; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D006801:Humans; D008156:Luciferases / Q000235:genetics; D008297:Male; D016328:NF-kappa B / Q000502:physiology*; D009362:Neoplasm Metastasis / Q000235:genetics / Q000473:pathology; D009363:Neoplasm Proteins / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011471:Prostatic Neoplasms / Q000235:genetics* / Q000473:pathology*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051785:Smad Proteins / Q000502:physiology*; D014158:Transcription, Genetic
C525718:BMP2 protein, human; C525809:BMP4 protein, human; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D016328:NF-kappa B; D009363:Neoplasm Proteins; D051785:Smad Proteins; D008156:Luciferases; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt